Part:BBa_K1632003

fim switch[default OFF](Tokyo_Tech/J23119)_rbs_gfp

The fim switch is the promoter containing repeated DNA sequence which is inverted by the Fim recombinase. Therefore, we can control the expression of the gene downstream of the fim switch by adding the Fim recombinase.
We designed this fim switch which has a J23119 promoter. Also between the promoter and the inverting site, there are two restriction enzyme sites in each front (SalIand BamHI) and back (BglII and MluI)(Fig. 1. fim switch (Tokyo_Tech/J23119) design). So the promoter can easily be interchanged. Except for insertion of restriction enzyme sites, basically, the design of fim switch(Tokyo_Tech) is similar with fim switch(wild-type).

Fig. 1. fim switch(Tokyo_Tech/J23119) design (Up:on state Down:off state)

From our results of our assay, the inversion of fim switch(Tokyo_Tech/J23119) by FimB/FimE was not confirmed correctly. The FimB protein inverts fim switch from [ON] state to [OFF] state and [OFF] state to [ON] state correctly (Fig.2.). However the FimE protein didn’t invert fim switch predominantly from [ON] state to [OFF] state. In the assay, the FimE protein inverts fim switch from [ON] state to [OFF] state and the [OFF] state to [ON] state (Fig.3.). In other words, the FimE protein works as the FimB protein.

Fig. 2. The histograms of the samples with FimB measured by flow cytometer

Fig. 3. The histograms of the samples with FimE measured by flow cytometer

For more information, see [http://2015.igem.org/Team:Tokyo_Tech/Project our work in Tokyo_Tech 2015 wiki].

More information

For more information, see http://2015.igem.org/Team:Tokyo_Tech/Project Our work in Tokyo_Tech 2015 wiki, http://2015.igem.org/Team:Tokyo_Tech/Experiment/ssrA_tag_degradation_assay About ssrA-tag, http://2015.igem.org/Team:Tokyo_Tech/Experiment/Overview_of_fim_inversion_system About ''fim'' inversion system

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 98
  Illegal NheI site found at 121
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 92
  Illegal BamHI site found at 133
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 1102